Expression and localization of epitope-tagged protein kinase CK2

✍ Scribed by C. Gail Penner; Zilong Wang; David W. Litchfield

• Publisher: John Wiley and Sons
• Year: 1997
• Tongue: English
• Weight: 355 KB
• Volume: 64
• Category: Article
• ISSN: 0730-2312
• DOI: 10.1002/(sici)1097-4644(19970315)64:4<525::aid-jcb1>3.0.co;2-t

✦ Synopsis

Protein kinase CK2, formerly known as casein kinase II, is a ubiquitous protein serine/threonine kinase. The enzyme exists in tetrameric complexes composed of two catalytic (CK2a and/or CK2a8) subunits and two subunits (CK2b) that appear to have a role in modulating the activity of the catalytic subunits. With the exception of their unrelated carboxy-terminal domains, the two isozymic forms of mammalian CK2 display extensive sequence identity. Furthermore, CK2a and CK2a8 exhibit remarkable conservation between species, suggesting that they may have unique functions. In the present study, the cDNAs encoding CK2a and CK2a8 were modified by addition of the hemagglutinin tag of the influenza virus at the amino terminus of the respective proteins. The epitope-tagged proteins were transfected into Cos-7 cells and the localization of the expressed proteins determined by indirect immunofluorescence using monoclonal antibodies specific for the epitope tag. The use of transfection favors the formation of homotetrameric complexes (i.e., a 2 b 2 , a8 2 b 2 ) instead of heterotetrameric complexes (i.e., aa8b 2 ) that are present in many cells. Epitope-tagged CK2a and CK2a8 displayed kinase activity and the ability to form complexes with CK2b. The results of these studies also indicate definitively that CK2a and CK2a8 are both localized predominantly within the nucleus. Mutation of conserved lysine residues within the ATP binding domains of CK2a and CK2a8 resulted in loss of kinase activity. However, examination of these mutants indicates that kinase activity is not essential for formation of complexes between subunits of CK2 and is not required for nuclear localization of CK2.